Abstract Fibrosis is a pathogenic process in organs (e.g., lung, kidney) involving the excess deposition of extracellular matrix (ECM) leading to loss of organ homeostasis. Fibrosis is the hallmark of progressive chronic kidney diseases as a common pathogenic response to injury. Similarly, end-stage lung diseases are often characterized by lung fibrosis. Recent studies suggest that necroptosis, a genetically-programmed form of cell death that is regulated by receptor-interacting protein-1 and -3 (RIPK1, RIPK3) kinases, may have emerging significance in human disease. Little is currently known of the role of RIPK3 in the pathogenesis of organ fibrosis. We have exciting preliminary data that RIPK3 can exert crucial functions in experimental models of kidney and lung fibrosis. Intriguingly, mice deficient in RIPK3, but not in its signaling target the mixed lineage kinase domain-like protein (MLKL), were protected against kidney fibrosis. We have also identified a RIPK3-mediated signaling pathway that regulates fatty acid (FA) metabolism by activating ATP citrate lyase (ACL), and contributes to kidney fibrosis. In contrast, mice deficient in either RIPK3 or MLKL were susceptible to pulmonary fibrosis. These studies suggest that RIPK3 may represent a novel mediator of organ fibrosis with differential organ or tissue-specific effects. The endogenous gaseous molecule carbon monoxide (CO) has been implicated as an experimental therapeutic modality in organ injury. Our published studies indicate that physiologic low-dose CO can mitigate fibrosis in unilateral ureteral obstruction (UUO)-induced kidney fibrosis, and in bleomycin (BLM)-induced pulmonary fibrosis. Therefore, we hypothesize that RIPK3 represents an important mediator of organ fibrosis through MLKL-independent and MLKL?dependent pathways. A RIPK3-dependent (MLKL-independent) signaling pathway and downstream regulation of the FA synthesis pathway contributes to the development of kidney fibrosis. In contrast, a RIPK3 and MLKL dependent pathway can inhibit pulmonary fibrosis. Moreover, we hypothesize that CO confers protection against multi-organ fibrosis by targeting either RIPK3 and/or FA- dependent pathways. RIPK3 and/or FA-biosynthetic proteins potentially serve as diagnostic biomarkers in predicting the severity of organ fibrosis and the efficacy of CO therapy. We will test these hypotheses in the following Specific Aims: Specific Aim 1: To characterize the function of RIPK3 and MLKL in the pathogenesis of organ fibrosis; Specific Aim 2: To determine the pathogenic contribution of RIPK3-regulated fatty acid (FA) synthesis in fibrotic organs; Specific Aim 3: To determine the role of the RIPK3 and the FA synthesis pathways in the therapeutic effects of CO in experimental lung and kidney fibrosis, and in human fibrosis.